Weidner D A, Winicov I
Department of Biochemistry University of Nevada Reno 89557.
Mol Cell Biochem. 1989 Oct 31;90(2):175-83. doi: 10.1007/BF00221217.
The beta-globin transcripts which are induced by dimethylsulfoxide (DMSO) and hexamethylene bisacetamide (HMBA) have been characterized in order to assess potential differences in their mechanisms of induction. Transcripts which initiate in the 5' flanking promoter region are likely indicators of promoter accessibility and were therefore characterized during the time course of induction with each inducer in Friend Erythroleukemia cells. S1 analysis with probes labeled at - 12 or +82 relative to the (+1) cap site showed no major differences between 5' ends of the upstream initiated transcripts in cells induced by DMSO or HMBA. We detected several upstream bands with each inducer corresponding to beta-globin transcripts with 5' ends between - 190 and -55 relative to the cap site and found that cells induced with DMSO and HMBA show a similar transcription response as measured by initiation in the 5' flanking region of the beta-globin gene. Interestingly, the upstream initiated transcripts reach their peak concentration levels much earlier in the time course of induction than do the mRNA transcripts with 5' ends at the major (+1) cap site. Northern blot analysis detected the upstream initiated transcripts as early as 16 hours after induction with DMSO, primarily in unprocessed large transcripts. We find that the promoter region containing transcripts constitute a higher percent of total beta-globin transcripts at the start of the induction and may therefore have an early function in the multistep induction process.
为了评估二甲亚砜(DMSO)和六亚甲基双乙酰胺(HMBA)诱导产生的β-珠蛋白转录本在诱导机制上的潜在差异,对其进行了表征。起始于5'侧翼启动子区域的转录本可能是启动子可及性的指标,因此在Friend红白血病细胞中用每种诱导剂进行诱导的时间进程中对其进行了表征。用相对于(+1)帽位点在-12或+82处标记的探针进行S1分析表明,在DMSO或HMBA诱导的细胞中,上游起始转录本的5'末端之间没有主要差异。我们用每种诱导剂检测到几个上游条带,它们对应于5'末端相对于帽位点在-190至-55之间的β-珠蛋白转录本,并且发现用DMSO和HMBA诱导的细胞在β-珠蛋白基因5'侧翼区域的起始转录方面显示出相似的转录反应。有趣的是,上游起始转录本在诱导时间进程中达到其峰值浓度水平的时间比5'末端在主要(+1)帽位点的mRNA转录本要早得多。Northern印迹分析早在用DMSO诱导后16小时就检测到了上游起始转录本,主要存在于未加工的大转录本中。我们发现,在诱导开始时,含有转录本的启动子区域在总β-珠蛋白转录本中所占百分比更高,因此可能在多步骤诱导过程中具有早期功能。